bims-mirnam Biomed News
on Mitochondrial RNA metabolism
Issue of 2025–11–30
fifteen papers selected by
Hana Antonicka, McGill University
  1. Transcriptomic analysis of cells following decreased mitochondrial DNA-copy number reveals compensatory mechanisms in mitochondrial DNA replication and cellular energetics.
  2. Electron microscopy visualization of cell-free mitochondrial DNA-containing extracellular vesicles in human plasma, serum, and saliva.
  3. Mitochondrial Base Editing of the m.8993T>G Mutation Restores Bioenergetics and Neural Differentiation in Patient iPSCs.
  4. Defining the impact of rRNA processing on nucleolar organization and function.
  5. DNASE1L3 surveils mitochondrial DNA on the surface of distinct mammalian cells.
  6. The proton motive force maintains mtDNA euploidy by balancing mtDNA replication with cell proliferation.
  7. Intrinsic errors in mitochondrial translation trigger a decline in cell fitness.
  8. Neuronal compartmentalization results in "impoverished" axonal mitochondria.
  9. KIF5A binds RNA to orchestrate synaptic mRNA localization and stress granules in ALS.
  10. mRNA caps accumulate in stress granules and are essential for their formation.
  11. Ribosome remodeling drives translation adaptation during viral infection and cellular stress.
  12. Nano-Mod-Amp reveals RNA sequence, structural and cell type specific features of pseudouridylation by PUS7.
  13. Long-read Sequencing of Nascent RNA from Budding and Fission Yeasts.
  14. Mutant p53 binds RNA to drive mitochondrial dysfunction.
  15. N6-methyladenosine regulation of mRNA translation is essential for early human erythropoiesis.
  1. bioRxiv. 2025 Nov 11. pii: 2025.11.10.687605. [Epub ahead of print]
    Transcriptomic analysis of cells following decreased mitochondrial DNA-copy number reveals compensatory mechanisms in mitochondrial DNA replication and cellular energetics.
    Jiaqi Xie, Phyo W Win, Charles Newcomb, Shaopeng Zeng, Christina A Castellani, Dan E Arking.
      Mitochondrial DNA copy number (mtDNA-CN) is a metric of mitochondrial function that has been associated with a variety of diseases including cardiovascular disease and all-cause mortality. To investigate genes and pathways affected by mtDNA-CN variation, we perturbed HEK 293T cells with ethidium bromide to deplete mtDNA. Using RNASeq and methylation microarrays, we evaluated transcriptomic and methylomic changes in treated cell lines. We observed an 8-fold decrease in mtDNA-CN and compensatory shifts in mitochondrial transcription to support mtDNA replication. Nuclear transcriptomic and methylomic analysis highlighted changes in metabolic pathways, including oxidative phosphorylation and canonical glycolysis. Longitudinal analyses revealed that the identified genes and pathways have different response timing, with nuclear response lagging behind mitochondrial response. These findings further elucidate the mechanisms behind mtDNA maintenance and responses to cellular energetics as well as mitochondrial-nuclear crosstalk dynamics.
    DOI:  https://doi.org/10.1101/2025.11.10.687605
  2. medRxiv. 2025 Oct 17. pii: 2025.10.15.25338094. [Epub ahead of print]
    Electron microscopy visualization of cell-free mitochondrial DNA-containing extracellular vesicles in human plasma, serum, and saliva.
    Alexandra Volos, Soah Grace Franklin, Jeremy Michelson, Shannon Rausser, Jonathan R Brestoff, Martin Picard.
      Human biofluids contain cell-free mitochondrial DNA (cf-mtDNA) and extracellular mitochondria (ex-Mito), creating the challenge of defining their origins, destinations, mechanisms of regulation, and purposes. To expand our understanding of cf-mtDNA biology, we present a descriptive electron microscopy analysis of circulating particles from cf-mtDNA-enriched plasma (citrate, heparin, and EDTA), serum (red and gold top), and saliva collected from ten healthy people (5 females, 5 males, mean age 44.9 years). Ex-mito and extracellular vesicles (EVs) were isolated by centrifugation followed by size-exclusion chromatography, imaged by transmission electron microscopy, and morphometrically analyzed. In parallel, cf-mtDNA was quantified in each biofluid. The resulting catalog of the most common circulating particles in plasma, serum, and saliva show that circulating double-membrane extracellular particles- consistent with mitochondrial ultrastructure-are present across human biofluids, along with EVs and other particle types. Combining imaging with cf-mtDNA quantification, we show that individuals with higher plasma cf-mtDNA concentrations tend to contain more double-membrane, ex-Mito-like particles. These preliminary results challenge the notion that, under normal conditions, the majority of cf-mtDNA exists as naked and potentially pro-inflammatory forms. Instead, these results are consistent with the concept of mitochondria transfer or signaling between cells and tissues. The image inventory provided here expands our knowledge of cell-free mitochondrial biology and provides a resource to inform biofluid selection and technical considerations in future studies quantifying ex-Mito and cf-mtDNA.
    DOI:  https://doi.org/10.1101/2025.10.15.25338094
  3. Genes (Basel). 2025 Nov 01. pii: 1298. [Epub ahead of print]16(11):
    Mitochondrial Base Editing of the m.8993T>G Mutation Restores Bioenergetics and Neural Differentiation in Patient iPSCs.
    Luke Yin, Angel Yin, Marjorie Jones.
       BACKGROUND: Point mutations in mitochondrial DNA (mtDNA) cause a range of neurometabolic disorders that currently have no curative treatments. The m.8993T>G mutation in the Homo sapiens MT-ATP6 gene leads to neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP) when heteroplasmy exceeds approximately 70%.
    METHODS: We engineered a split DddA-derived cytosine base editor (DdCBE), each half fused to programmable TALE DNA-binding domains and a mitochondrial targeting sequence, to correct the m.8993T>G mutation in patient-derived induced pluripotent stem cells (iPSCs). Seven days after plasmid delivery, deep amplicon sequencing showed 35 ± 3% on-target C•G→T•A conversion at position 8993, reducing mutant heteroplasmy from 80 ± 2% to 45 ± 3% with less than 0.5% editing at ten predicted off-target loci.
    RESULTS: Edited cells exhibited a 25% increase in basal oxygen consumption rate, a 50% improvement in ATP-linked respiration, and a 2.3-fold restoration of ATP synthase activity. Directed neural differentiation yielded 85 ± 2% Nestin-positive progenitors compared to 60 ± 2% in unedited controls.
    CONCLUSIONS: Edits remained stable over 30 days in culture. These results establish mitochondrial base editing as a precise and durable strategy to ameliorate biochemical and cellular defects in NARP patient cells.
    Keywords:  DdCBE; MT-ATP6; NARP; base editing; heteroplasmy; iPSCs; m.8993T>G; mitochondrial DNA
    DOI:  https://doi.org/10.3390/genes16111298
  4. bioRxiv. 2025 Oct 19. pii: 2025.10.19.682850. [Epub ahead of print]
    Defining the impact of rRNA processing on nucleolar organization and function.
    Maria Saraí Mendoza-Figueroa, Ellen Lavorando, Yulia Gonskikh, Charles Antony, Heidi Elashal, Anthony Yulin Chen, Hsin-Yao Tang, Dawn Carone, Vikram R Paralkar, Kathy Fange Liu.
      The eukaryotic nucleolus is a highly organized, multilayered structure essential for ribosomal RNA (rRNA) processing and ribosome assembly. However, how the sequential steps of rRNA maturation, particularly the series of endonucleolytic cleavages, contribute to maintaining nucleolar architecture remains poorly understood. Here, we show that disruption of pre-rRNA processing, especially impaired cleavage of the 5' external transcribed spacer (5'ETS), profoundly alters nucleolar organization. Specifically, defects in 5'ETS processing lead to the formation of a single large DAPI-negative nuclear structure and result in the mislocalization of nascent RNA, which diffuses throughout the disorganized nucleolus. These aberrant nucleoli exhibit a distinct proteomic profile, including downregulation of factors involved in splicing, cell cycle regulation, and chromatin organization, suggesting that the impact of nucleolar disorganization extends beyond ribosome biogenesis. Notably, we also observe mislocalization of heterochromatin markers, pointing to broader disruptions in nuclear architecture and gene regulation. Together, our findings reveal that proper 5'ETS cleavage is critical for preserving nucleolar compartmentalization and highlight the tight coupling between rRNA processing and nuclear organization.
    DOI:  https://doi.org/10.1101/2025.10.19.682850
  5. bioRxiv. 2025 Oct 06. pii: 2025.10.05.680464. [Epub ahead of print]
    DNASE1L3 surveils mitochondrial DNA on the surface of distinct mammalian cells.
    Jennifer Porat, Valentina Poli, Karim Almahayni, Benson M George, Francisca Nathália de Luna Vitorino, Li Yi, Carolina Bras Costa, Yitong Zhou, Andrew Crompton, Hongxiu Wen, Leon Zheng, Peter K Gregersen, Suneet Agarwal, Benjamin A Garcia, Yixuan Xie, Leonhard Möckl, Ivan Zanoni, Kim R Simpfendorfer, Ryan A Flynn.
      The extracellular space is a critical environment for discriminating self versus non-self nucleic acids and initiating the appropriate immune responses through signaling cascades to relay information about extracellular nucleic acids. Here, we provide evidence that oxidized mitochondrial DNA is tethered to the surface of select mammalian cells through cell surface proteins and heparan sulfate proteoglycans. We demonstrate that cell surface DNA accumulates in large clusters that partially overlap with domains enriched in RNA binding proteins. Finally, we show that human and murine B cell surfaces contain DNA that can be cleared by the secreted nuclease DNASE1L3, and that patients with a DNASE1L3 missense variant associated with increased risk for autoimmune disease harbor increased levels of surface DNA on B and T cells. Taken together, this work expands the scope of cell surface nucleic acid biology and provides a mechanistic link between cell surface molecules and DNA targeting in autoimmune disease.
    DOI:  https://doi.org/10.1101/2025.10.05.680464
  6. bioRxiv. 2025 Oct 27. pii: 2025.10.27.684790. [Epub ahead of print]
    The proton motive force maintains mtDNA euploidy by balancing mtDNA replication with cell proliferation.
    Sneha P Rath, Vamsi K Mootha.
      Human mitochondrial DNA (mtDNA) encodes 13 essential components of the electron transport chain (ETC) 1 . A typical cell contains ∼1000s of copies of mtDNA, but how this copy number is stably maintained is unclear. Here, we track mtDNA copy number (mtCN) recovery in K562 cells following transient, chemically induced depletion to uncover principles of mtCN stability. Below a critical mtCN, ETC activity fails to sustain the proton motive force (PMF) and de novo pyrimidine synthesis-both required for mtDNA replication. PMF-dependent processes like Fe-S cluster biogenesis are also disrupted and stress responses are activated that impair cell proliferation and limit further mtCN dilution by cell division. Nonetheless, mtDNA replication and recovery remain possible via mtDNA-independent PMF, generated by complex V reversal, and uridine salvage. Once mtCN is restored, the ETC and forward complex V activity re-engage, stress responses subside, and proliferation recommences. Each cell division then dilutes mtDNA, serving as a built- in brake on mtCN. Our findings suggest that mtCN homeostasis emerges from the balance of two opposing PMF-driven processes - mtDNA replication and cell proliferation - revealing a bioenergetic logic that preserves mtDNA euploidy through repeated cell divisions.
    DOI:  https://doi.org/10.1101/2025.10.27.684790
  7. bioRxiv. 2025 Oct 15. pii: 2025.10.13.682189. [Epub ahead of print]
    Intrinsic errors in mitochondrial translation trigger a decline in cell fitness.
    Güleycan Lutfullahoglu Bal, Kah Ying Ng, Eli Berzell, Ani Akpinar, Alex E Ekvik, Lidiia Koludarova, Seyedehshima Naddafi, Clemens Raths, Tania Vane-Tempest, Jordyn J VanPortfliet, Sarah O'Keefe, Arafath K Najumudeen, Tuula A Nyman, James B Stewart, A Phillip West, Brendan J Battersby.
      Defects in the faithful expression of the human mitochondrial genome underlies disease states, from rare inherited disorders to common pathologies and the aging process itself. The ensuing decrease in the capacity for oxidative phosphorylation alone cannot account for the phenotype complexity associated with disease. Here, we address how aberrations in mitochondrial nascent chain synthesis per se exert a decline in cell fitness using a classic model of mitochondrial induced premature aging. We identify how intrinsic errors during mitochondrial nascent chain synthesis destabilize organelle gene expression, triggering intracellular stress responses that rewire cellular metabolism and cytokine secretion. Further, we show how these mechanisms extend to pathogenic variants associated with inherited human disorders. Together, our findings reveal how aberrations in mitochondrial protein synthesis can sensitize a cell to metabolic challenges associated with disease and pathogen infection independent of oxidative phosphorylation.
    Teaser/One-Sentence Summary: Aberrations in mitochondrial translation elongation trigger activation of intracellular stress responses associated with disease and aging.
    DOI:  https://doi.org/10.1101/2025.10.13.682189
  8. bioRxiv. 2025 Oct 29. pii: 2025.10.27.684882. [Epub ahead of print]
    Neuronal compartmentalization results in "impoverished" axonal mitochondria.
    Adrian Marti Pastor, Liam S C Lewis, Stephan Müller, Pedro Guedes-Dias, Lorena Olifiers, Polina Pelkonen, Almudena Del Rio Martin, Caroline Fecher, Nikolai P Skiba, Ying Hao, Antoneta Gavoci, Laura Trovo, Mehmet Akif Aktas, Hariharan M Mahadevan, Angelika B Harbauer, Howard M Bomze, Vadim Y Arshavsky, Monika S Brill, Romain Cartoni, Stefan F Lichtenthaler, Sidney M Gospe, Thomas Misgeld.
      Mitochondria differ depending on their location within a neuron. Morphological heterogeneity between somatic, dendritic, and axonal mitochondria is well established. Emerging evidence suggests that further specialization is needed to meet the unique demands of different neuronal compartments. However, the molecular and functional diversity of mitochondria within a neuron remains poorly understood. Here, we utilized proteomics in MitoTag mice to profile somatodendritic and axonal mitochondria across four distinct neuron types, thereby generating a compendium of intracellular mitochondrial diversity. Combining proteomics, functional, and immunofluorescence analyses, we demonstrated that axonal mitochondria are not defined by the presence of unique proteins, but rather by the selective loss or preservation of specific pathways compared to their somatodendritic counterparts. This results in "impoverished" axonal mitochondria, which are characterized by diminished mtDNA expression and impaired oxidative phosphorylation yet retain other pathways, such as fatty acid metabolism. Bioinformatic analyses of multiomic data identified local translation as one mechanism underlying compartment-specific diversity. Together, these findings provide a comprehensive in vivo framework for understanding mitochondrial specialization across neuronal compartments.
    DOI:  https://doi.org/10.1101/2025.10.27.684882
  9. bioRxiv. 2025 Nov 02. pii: 2025.10.31.685813. [Epub ahead of print]
    KIF5A binds RNA to orchestrate synaptic mRNA localization and stress granules in ALS.
    Phuong Le, Neeraj K Lal, Shuhao Xu, Sara Mumford, Megan Huang, Dong Yang, Orel Mizrahi, Benjamin Hoover, Brian Yee, Yuan Mei, Katherine Rothamel, Hsuan-Lin Her, Steven M Blue, Neil A Shneider, Gene W Yeo.
      Neuronal health depends on the precise transport and local translation of mRNAs to maintain synaptic function across highly polarized cellular architecture. While kinesin motor proteins are known to mediate mRNA transport, the specificity and direct involvement of individual kinesins as RNA-binding proteins (RBPs) remain unclear. Here, we demonstrate that KIF5A, a neuron-specific kinesin implicated in amyotrophic lateral sclerosis (ALS), functions as an RBP. We show that KIF5A directly binds mRNAs encoding synaptic ribosomal proteins and is required for their synaptic localization and for maintaining normal synaptic composition and function. Additionally, we show ALS-linked KIF5A mutations confer gain-of-function properties, enhancing mRNA binding, increasing synaptic ribosomal protein accumulation, inducing neuronal hyperexcitability, and impairing stress responses. These findings reveal a previously unrecognized mechanism by which mutant KIF5A disrupts synaptic homeostasis. Our work positions a kinesin motor protein as an RBP with critical roles in mRNA transport, local translation, and stress response.
    Highlights: KIF5A interacts with mRNA encoding synaptic ribosomal proteinsKIF5A is required for normal synaptic composition and functionKIF5A binds to G3BP1 and G3BP1 stress granule associated proteinsKIF5A mutant ALS patient-derived motor neurons have abnormal synaptic function and stress response.
    DOI:  https://doi.org/10.1101/2025.10.31.685813
  10. Cell Mol Life Sci. 2025 Nov 25. 82(1): 420
    mRNA caps accumulate in stress granules and are essential for their formation.
    Mingxin Guo, Ying Feng, Guantong Qi, Wenran Ma, Xiaoli He, Ruiqi Li, Yu Liu, Jin Wang.
      Stress granules (SGs) are essential cytoplasmic, membraneless organelles that form in response to cellular stress, functioning to prevent mRNA translation and protect mRNA from damage. However, the mechanism of SG formation remains largely unknown. Here, utilizing a systems-level technique for quantification of RNA cap epitranscriptome, we found that mRNA Cap1 and non-canonical caps are predominantly enriched in SGs, with the composition of mRNA caps in SGs of mammalian cells differing between different stress conditions. Knockdown of RNA guanine-7-methyltransferase (RNMT) and phosphorylated CTD interacting factor 1(PCIF1) both resulted in substantial changes in the content and composition of mRNA caps and RNMT knockdown caused failure of SG formation under different stress conditions. Furthermore, proteomic, Co-IP and confocal immunofluorescence analysis of these knockdown cells and SGs revealed that mRNAs partition into SGs through cap-protein interactions. These findings collectively revealed the significant role of mRNA cap in SG formation and stress response in mammalian cells.
    Keywords:  CapQuant; Stress; Stress granule; mRNA cap
    DOI:  https://doi.org/10.1007/s00018-025-05947-8
  11. bioRxiv. 2025 Oct 24. pii: 2025.10.24.684008. [Epub ahead of print]
    Ribosome remodeling drives translation adaptation during viral infection and cellular stress.
    Hsin-Yu Tsai, Luochen Liu, Rebecca H Fleming, Julian Mintseris, Derrick Ekanayake, Xin Gu, Steven P Gygi, Amy S Y Lee.
      The ribosome is the highly conserved molecular machine that decodes mRNAs during protein synthesis. While traditionally thought to consist of a uniform set of proteins, here we discover that ribosome composition is reprogrammed to adapt to intrinsic and external cellular perturbations. During infection by non-segmented negative-sense viruses, viral entry into cells recruits the large ribosomal subunit protein rpL40 to a noncanonical site on the small subunit of 80S ribosomes near the mRNA entry site. These specialized ribosomes preferentially bind viral mRNAs to drive enhanced viral protein synthesis that is critical for replication under host pressures. Unexpectedly, we find that viruses have co-opted this translation pathway from a previously unrecognized endogenous ribosome remodeling program in which metabolic stress alters ribosome structure to promote mRNA translation required for cell survival. Thus, ribosome remodeling is a conserved mechanism enabling dynamic protein synthesis across pathogen and cellular adaptation.
    DOI:  https://doi.org/10.1101/2025.10.24.684008
  12. bioRxiv. 2025 Oct 31. pii: 2025.10.30.685621. [Epub ahead of print]
    Nano-Mod-Amp reveals RNA sequence, structural and cell type specific features of pseudouridylation by PUS7.
    Rebecca Rodell, Ronit Jain, Hossein Shenasa, Matias Montes, Nicolas Robalin, Stefan Prodic, Eduardo Eyras, Nicole M Martinez.
      Pseudouridines are abundant mRNA modifications that can impact splicing, translation, and stability to tune gene expression. PUS7 is one of the major mRNA pseudouridine synthase whose dysregulation leads to neurodevelopmental disorders and cancer, underscoring the critical function of PUS7-dependent pseudouridines. Beyond a short and degenerate consensus sequence, the molecular mechanisms underlying PUS7-mediated pseudouridylation remain unknown. A lack of targeted, high-throughput pseudouridine detection methods limits simultaneous interrogation of PUS7 regulatory features across many experimental conditions. We developed novel Nanopore sequencing tools, including Nano-Mod-Amp, to reveal pseudouridine stoichiometry, its RNA structural context, and dependence on PUS7 levels at specific sites across biological conditions. We identified a novel RNA structural signature that is associated with more efficient mRNA modification by PUS7. Pseudouridines are largely responsive to modulations in PUS7 protein levels, demonstrating the regulatory potential of varying PUS7 levels across cellular conditions. Conversely, PUS7 activity is also regulated in a cell-type specific manner, independent of PUS7 expression levels in a manner consistent with regulation by RNA structure and RNA binding proteins. Together, we developed Nanopore sequencing tools and uncovered new mechanisms of PUS7 regulation with a framework that can be applied to other RNA-modifying enzymes to query the regulation of the epitranscriptome.
    Highlights: Nanopore direct RNA sequencing identifies PUS7-dependent pseudouridines with stoichiometry.Nano-Mod-Amp quantifies PUS7-dependent pseudouridines at hundreds of sites in high-throughput.MPRAs define RNA sequence and structural features associated with modification by PUS7.Individual PUS7 target pseudouridines are substoichiometric and poised for regulation.PUS7 activity is regulated by cell type in the absence of differences in PUS7 protein levels.
    DOI:  https://doi.org/10.1101/2025.10.30.685621
  13. bioRxiv. 2025 Oct 06. pii: 2025.10.06.680282. [Epub ahead of print]
    Long-read Sequencing of Nascent RNA from Budding and Fission Yeasts.
    Kyle D Robik, Tara Alpert, Kirsten A Reimer, Pernille Bech, Lydia Herzel, Leonard Schärfen, Korinna Straube, Karla M Neugebauer.
      Gene expression requires DNA transcription and simultaneous RNA processing steps that transform the precursor RNA into fully mature RNA. In eukaryotes, the processing of protein-encoding messenger RNAs (mRNAs) includes 5' end capping, editing, splicing, RNA modification, poly-adenylation cleavage, and polyadenylation. Short-read sequencing of total or messenger RNA largely reveals the final output of transcription and processing because it utilizes 1) steady-state, mature RNA that is mostly processed and 2) sequencing reads that are too short to detect adjacent processing events (e.g. two adjacent introns). In contrast, long-read sequencing of nascent RNA allows the detection of rarer, full-length transcripts that are in the process of being transcribed and processed. The 3' end of each nascent RNA establishes the position of RNA polymerase II (Pol II) along the gene at the time of cell lysis, providing a 'timeline' for RNA processing events. In addition, the density of 3' ends along genes or at gene landmarks reflects Pol II density, which is related to changes in transcription elongation rate. In organisms with complex gene architectures, information about splicing across multiple introns within the same transcript can be extracted, as well as the location of transcription start sites (TSSs) and polyA cleavage sites. Here, we describe the isolation of nascent RNA from the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe , preparation of a cDNA library for long-read sequencing on Oxford Nanopore Technologies or Pacific Biosciences platforms, and initial data analysis steps. These methods comprise versatile and powerful tools for the investigation of coupled RNA synthesis and processing.
    DOI:  https://doi.org/10.1101/2025.10.06.680282
  14. bioRxiv. 2025 Nov 13. pii: 2025.11.12.688127. [Epub ahead of print]
    Mutant p53 binds RNA to drive mitochondrial dysfunction.
    Wuyue Zhou, Alice Long, Cameron J Douglas, Dalila Gallegos, James M Burke, David W C MacMillan, Ezgi Hacisuleyman, Ciaran P Seath.
      Tumor suppressor protein 53 (p53) is a transcription factor that is deregulated in 50% of cancers. Often termed the guardian of the genome, p53 is responsible for maintenance of genomic stability, cell cycle arrest, DNA repair, senescence, and apoptosis. In cancer cells, deregulation of p53 often occurs through mutations in the DNA binding domain which lead to a loss of the transcriptional activity. While 100's of somatic mutations in the DNA binding domain are known, a small number of mutants are enriched in cancer, suggesting a gain-of-function role. Here we deploy an intein-based approach to localize µMap photoproximity labeling to p53 to define novel interactions contributing to the loss and gain of function roles of 5 separate hotspot mutants. These data revealed that G245S and R273H binds to RNA through its C-terminal domain. We show through CLIP experiments that mutant p53 has an RNA binding motif that conserved across mutants and is enriched in 3'UTRs, promoting ribosomal localization and labeling of proteins at the mitochondrial surface. We further demonstrate that the RNA binding ability of mutant p53 promotes altered miRNA processing and mitochondrial dysfunction providing mechanistic rationale for historically reported but poorly understood phenotypes.
    DOI:  https://doi.org/10.1101/2025.11.12.688127
  15. bioRxiv. 2025 Nov 12. pii: 2025.11.10.687731. [Epub ahead of print]
    N6-methyladenosine regulation of mRNA translation is essential for early human erythropoiesis.
    Daniel A Kuppers, Sonali Arora, Cindy L Wladyka, Ruiqi Ge, Shun Liu, Yong Peng, Rui Su, Anne Wilhite, Jianjun Chen, Chuan He, Andrew C Hsieh, Patrick J Paddison.
      N6-methyladenosine (m 6 A) is an abundant modification of mRNA with important regulatory roles in normal and malignant hematopoiesis. We previously reported that in human erythroid leukemia (HEL) cells, m 6 A mRNA marking selectively regulates translation of essential erythropoiesis genes required for in vitro differentiation and human erythroid colony formation. Here, we further investigated the timing and nature of requirement for m 6 A-methyltransferase (MTase) activity during human erythropoiesis, using a standardized in vitro erythroid differentiation assay for hHSPCs. We identified two critical m 6 A regulated developmental windows in BFU-E and during the transition from CFU-E to proerythroblasts. These windows of m 6 A-MTase requirement coincide with rising global m 6 A levels, which peak in proerythroblasts. After proerythroblast formation, however, m 6 A -MTase activity is dispensable for differentiation, proliferation, and survival. In BFU-E, m 6 A-MTase promotes proliferation but is dispensable for differentiation, while, in CFU-E, both m 6 A -MTase and the YTHDF family of m 6 A readers are essential for differentiation to proerythroblasts. Mechanistically, in CFU-E, m 6 A MTase activity enhances translation of ribosomal and oxidative phosphorylation (OXPHOS) genes, thereby elevating global protein synthesis rates and enabling efficient erythroblast formation. We propose that this form of translational regulation by m 6 A emerged as an evolutionary adaptation to meet the high translational demands of human erythropoiesis.
    DOI:  https://doi.org/10.1101/2025.11.10.687731
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